The overall theme of our current proposal is to delineate protocols for the induction and maintenance of donor specific tolerance in recipients of islet allografts, with our ultimate goal being the reversal of insulin dependent diabetes mellitus. We plan to thoroughly investigate three approaches which have yielded promising results in animal models: l)intrathymic inoculation of donor cells and/or antigen, primarily to mediate clonal deletion of anti-donor specific T cells, and 2) intravenous administration of donor bone marrow cells, with the aim of inducing a state of microchimerism and/or recruiting both central and peripheral mechanisms of tolerance, 3) inhibition of costimulatory (B7/CD28/CTLA-4) and adhesive interactions between recipient leukocytes and donor cells or tissues. We will define the mechanism(s) of donor specific tolerance by evaluating in vitro and in vivo immunoreactivity to both donor and 3rd party cells and tissues, and in addition, will search for the presence of chimerism in bone marrow/islet recipients. We will also explore the variables inherent in each protocol by analyzing the type and timing of immunosuppression relative to intrathymic inoculation or i.v. bone marrow infusion. Interference in such pathways, i.e., recognition of donor alloantigen in the absence of the appropriate costimulatory signals, can lead to clonal anergy. Findings from these studies have the potential of directing us towards a multifaceted approach to the induction of donor specific tolerance in humans. In addition, the induction of tolerance to transplanted islets may provide the added benefit of tolerizing a diabetic patient to the grafted islets, thus preventing recurrence of the autoimmune process. The availability of the findings from these studies will provide key insights that will be applied to the refinement of ongoing clinical trials of islet allotransplantation in humans. The application of this information should, as well, be generally applicable to the field of organ transplantation.